1. Field of the Invention
The present invention relates to a new combined preparation for the treatment of neoplasic diseases or of infectious diseases.
The present invention describes sequences of conventional treatments of cancer or infections and of immunotherapies reversing or preventing chemoresistance and allowing long lasting therapeutic responses.
2. Description of the Related Art
In conventional therapy, residual tumor cells or infectious agents are left undamaged due to chemoresistance or due to the fact that these cells are shaded in protected areas or located in hypoxic areas poorly vascularized and not accessible to conventional treatments. The genetic instability and heterogeneity of tumors and micro-organisms indeed allow them to adapt and to develop resistance to therapies.
The beneficial effects of chemotherapy can be compromised by cellular mechanisms that allow infectious agents or neoplasic tissue to evade the toxicity of drugs. In some cases, pleiotropic resistance to a variety of unrelated drugs has been observed, and this phenomenon has been called multidrug resistance.
Resistance to chemotherapy, whether it is intrinsic or acquired, is a major cause of failure in the curative treatment of chronic infections or neoplasic malignancies. Among the most active anti-cancer agents used in the treatment of haematological malignancies are some natural toxin-derived drugs, such as the anthracycline daunorubicin or adriamicin, the epipodophyllotoxins, taxoter derivatives, the vinca alkaloid vincristine, cisplatin, fluorouracils.
Development of cross-resistance to these structurally and functionally unrelated drugs, called multidrug resistance, is frequently observed in second or third intention cytotoxic treatment of cancer.
Multiple drug resistance of infectious agents and particularly of bacteria to antibiotics such as penicilins, xcex2-lactamines, cephalosporines, aminoglucosides, macrolides and sulfamides, is more and more often seen in hospitals.
Monocytes derived cells (MDCs) are immune cells such as obtained by culture of blood mononuclear cells in non adherent gas permeable plastic or Teflon bags for 5 to 10 days at 37xc2x0 C. in O2/CO2 atmosphere. Their culture medium (RPMI, IMDM, AIM5 (Gibco) or X-VIVO (Biowhittaker)) contains eventually cytokines or ligands as defined in patents no PCT/EP93/01232, no WO94/26875 or EP 97/02703 or in the articles mentioned below:
xe2x80x9cAutologous lymphocytes prevent the death of monocytes in culture and promote, as do GM-CSF, IL-3 and M-CSF, their differentiation into macrophagesxe2x80x9d. (Lopez M., Martinache Ch., Canepa S., Chokri M., Scotto F., Bartholeyns J.; J. of Immunological Methods, 159: 29-38, 1993);
xe2x80x9cImmnune therapy with macrophages: Present status and critical requirements for implementationxe2x80x9d (Bartholeyns J., Romet-Lemonne J-L., Chokri M., Lopez M.; Immunobiol., 195: 550-562, 1996);
xe2x80x9cIn vitro generation of CD83+ human blood dendritic cells for active tumor immunotherapyxe2x80x9d (Thurnher M., Papesh C., Ramoner R., Gastlt G. and al.; Experimental Hematology, 25: 232-237, 1997);
xe2x80x9cDendritic cells as adjuvants for immnune-mediated resistance to tumorsxe2x80x9d (Schuler G. and Steinman R. M.; J. Exp. Med., 186: 1183-1187, 1997).
All these patents applications and articles are included herein for references.
They can be activated by INF-xcex3 at the end of culture to obtain in particular cytotoxic macrophages. They can be centrifuged to be concentrated and purified before resuspension in isotonic solution.
Monocytes derived cells (MDCs) can either be killer macrophages, phagocytozing cells, growth factors and cytokines releasing cells, or dendritic cells according to their conditions of differentiation. Dendritic cells can for example be obtained as described in xe2x80x9cIn vitro generation of CD83+ human blood dendritic cells for active tumor immunotherapyxe2x80x9d (Thurnher M., Papesh C., Ramoner R., Gastlt G. and al.; Experimental Hematology, 25: 232-237, 1997) and xe2x80x9cDendritic cells as adjuvants for immune-mediated resistance to tumorsxe2x80x9d (Schuler G. and Steinman R. M.; J. Exp. Med., 186: 1183-1187, 1997), and EP no 97/02703.
In addition, activated monocyte derived cells (macrophages) can be used to deliver therapeutic agents to tumor or infectious sites.
One of the aims of the invention is to provide a combined preparation of active substances under the form of individual components for the simultaneous separate or sequential use, in the treatment of cancer or of infectious diseases.
Another aim of the invention is to provide a method for the treatment of residual cancer resistant to chemotherapy.
Another aim of the invention is to provide a method for the treatment of infectious diseases resistant to antibiotic treatment.
The invention relates to a combined preparation containing, as active substance, the following individual components, in the form of a kit-of-parts:
monocyte derived cells, particularly cytotoxic macrophages,
chemotherapy or immunotherapy drugs,
for the simultaneous, separate or sequential use, for the treatment of cancer or infectious diseases.
The present treatment consists in the local or systemic injection of autologous activated macrophages (MAK(copyright) killer cells) or monocyte derived cells which have access to injured areas, and in particular to hypoxic areas, where they tend to concentrate.
This treatment can be conducted after first failure and relapse following chemotherapies, or before chemotherapy, to prevent chemoresistance. Local treatment with chemotherapy drugs causes cell necrosis and release of chemokines which call and actively recrute macrophages and monocytes derived cells. Therefore, combining the chemotherapy with macrophage immunotherapy can in synergy increase cytotoxicity and increase immune response at the same time as preventing the establishment of resistance. Additionally to a first treatment combining conventional approach with immunotherapy, macrophage adoptive therapy can be proposed after failure and relapse.
It is shown through the invention that the local or systemic injection of activated monocyte derived cells, or macrophages, restores clinical responses to cytotoxic drugs for which resistance was previously demonstrated, or prevents the apparition of chemoresistance.
The present invention also shows that activated monocyte derived cells can overcome this resistance and synergize for therapy.
The two active ingredients of the combined preparation have never been used for a new joint effect and are unknown as a composition.
The active ingredients which are administered either at the same time, or separately, or sequentially, according to the invention, do not represent a mere aggregate of known agents, but a new combination with the surprising valuable property that immunotherapy with monocyte derived cells modifies the chemoresistance/chemosensitivity and allows a new effective treatment (partial or complete response) with similar chemotherapy protocole. Furthermore, synergy is observed between monocyte derived cells immunotherapy and chemotherapy.
It is to be stressed that the combined preparation also designated by a kit-of-parts means that the components of the combined preparation are not necessarily present as a union e.g. in composition, in order to be available for separate or sequential application. Thus the expression kit-of-parts means that it is not necessarily a true combination, in view of the physical separation of the components.
In an advantageous combined preparation of the invention, the monocyte derived cells contain chemotherapy or immunotherapy drugs.
In another advantageous combined preparation of the invention, the monocyte derived cells are such as prepared according to the method comprising the following steps:
1) recovery of blood derived mononuclear cells directly from blood apheresis or from blood bag collection, followed if necessary by centrifugation, to eliminate a substantial part of red blood cells, granulocytes and platelets, and collection of peripheral blood leukocytes;
2) washing peripheral blood leukocytes obtained at the preceeding steps for instance by centrifugation (to remove 90% of platelets, red blood cells and debris) to obtain mononuclear cells;
3) resuspension of the total mononuclear cells (monocytes+lymphocytes) obtained at the preceeding step in culture medium (RPMI or IMDM type) at 106 to 2.107 cells/ml, possibly completed by cytokines and/or autologous serum, and culture for 5 to 10 days at 37xc2x0 C. under O2/CO2 atmosphere in hydrophobic gas permeable bags, to obtain monocyte derived cells and contaminating lymphocytes.
According to an advantageous combined preparation, the chemotherapy drug is selected among cytotoxic compounds such as anthracyclins, daunorubicin, adriamycin, taxoter derivatives, vinca alcaloids, vincristine, carmustine, cisplatin, fluorouracils, cytostatic compounds such as polyamine inhibitors, topoisomerase inhibitors, tamoxifene, prodasone, or sandostatine, or compounds inducing apoptosis such as sodium butyrate or mitomycin C, antibiotics such as penicilins, xcex2-lactamines, cephalosporines, cyclines, aminoglucosides, macrolides or sulfamides, or antiviral drugs such as AZT, protease inhibitors or acyclovir, retrovir or foscarnet.
According to an advantageous embodiment, the combined preparation of the invention is such that the immunotherapy drug is selected among cytokines such as cyclosporine, azathioprine, cyclophosphamide, IFNxcex3, IL-12, IL-2, GM-CSF, G-CSF, and immunoadjuvants such as murapeptides or BCG.
According to an advantageous embodiment, in the combined preparation of the invention, the monocyte derived cells and the chemotherapy or immunotherapy drugs are in the form in injectable solutions.
In another advantageous embodiment of the invention, in the combined preparation, the injectable solutions are in the form of locally injectable solutions.
In another advantageous embodiment in the combined preparation of the invention, the injectable solutions are in the form of systemically injectable solutions.
In another advantageous combined preparation of the invention, the monocyte derived cells are administered at a dose of about 107 to about 1010 monocyte derived cells per injection.
In another advantageous combined preparation of the invention, the monocyte derived cells are administrated at a dose of about 108 to about 109. 
In another advantageous combined preparation of the invention, the monocyte derived cells are administered in a repeated way up to ten times, the interval between each administration being between three days to two months.
In another advantageous combined preparation of the invention, the immunotherapy or chemotherapy drug is administered at a dose of about 0.1 to about 1000 mg/day.
In another advantageous combined preparation of the invention, in the case of administration of a drug chosen among immunotherapy drug, antiviral drug, cytotoxic drugs, or antibiotics, said drug is administered at a dose of about 10 to about 1000 mg/day.
More specifically, in the case of cytotoxic compounds such as vincristine, taxol, carmustine, daunorubicin, adryamicin, cisplatin, fluorouracil, they are administered at a dose of about 10 to about 500 mg/day.
In the case of antiviral drugs such as retrovir, aciclovir, foscarnet, said drug is administered at a dose of about 20 to about 500 mg/day.
In the case of antibiotics such as penicilins, cephalosporine, sulfamides, cyclines, said drug is administered at a dose of about 10 to about 1000 mg/day.
In the case of immunotherapy drugs such as cyclosporine, azathioprine, cyclophosphamide, said drug is administered at a dose of about 10 to about 1000 mg/day.
In another advantageous combined preparation of the invention, in the case of administration of a drug chosen among cytostatic compounds, apoptosis inducing compounds or cytokines, said drug is administered at a dose of about 0.1 to about 100 mg/day.
In the case of cytostatic compounds such as amoxifene, prodasone, sandostatine, polyamine inhibitors or apoptosis inducing compounds such as sodium butyrate or mitomycin C, said drug is administered at a dose of about 0.1 to about 100 mg/day.
In another advantageous combined preparation of the invention, the immunotherapy or chemotherapy drug is administered in a repeated way up to 10 times, the interval between each administration being between one day to two months.
In another advantageous combined preparation of the invention, the chemotherapy or immunotherapy drug and the monocyte derived cells are injected simultaneously.
In another advantageous combined preparation of the invention, the chemotherapy or immunotherapy drug and the monocyte derived cells are administered in sequential way, the immunotherapy or chemotherapy drug being administered before the monocyte derived cells.
In another advantageous combined preparation of the invention, the interval of time between the administration of the monocyte derived cells and the administration of the immunotherapy or chemotherapy drugs is of one day to two months.
In another advantageous combined preparation of the invention, the monocyte derived cells and the chemotherapy or immunotherapy drug are administered seqentially, the monocytes derived cells being administered before the immunotherapy or chemotherapy drug.
In another advantageous combined preparation of the invention, the interval of time between the administration of the immunotherapy or chemotherapy drug and the administration of the monocyte derived cells is of one day to two months.
In another advantageous combined preparation of the invention, the administration of monocyte derived cells is followed by an administration of chemotherapy or immunotherapy drug.
In another advantageous combined preparation of the invention, the interval of time between the administration of monocyte derived cells and the administration of chemotherapy or immunotherapy drugs is of one day to two months.
The invention also relates to a method for the treatment of residual cancer resistant to chemotherapy or of infectious diseases resistant to chemotherapy comprising the use of a combined preparation of the invention.
The invention also relates to a method for the treatment of infectious diseases resistant to antibiotic treatment comprising the use of a combined preparation of the invention.